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Showing papers in "Lab on a Chip in 2014"


Journal ArticleDOI
TL;DR: A three dimensional micromolding technique utilizing bioprinted agarose template fibers to fabricate microchannel networks with various architectural features within photocrosslinkable hydrogel constructs with useful applications in tissue engineering and organs on a chip is reported.
Abstract: Vascularization remains a critical challenge in tissue engineering. The development of vascular networks within densely populated and metabolically functional tissues facilitate transport of nutrients and removal of waste products, thus preserving cellular viability over a long period of time. Despite tremendous progress in fabricating complex tissue constructs in the past few years, approaches for controlled vascularization within hydrogel based engineered tissue constructs have remained limited. Here, we report a three dimensional (3D) micromolding technique utilizing bioprinted agarose template fibers to fabricate microchannel networks with various architectural features within photocrosslinkable hydrogel constructs. Using the proposed approach, we were able to successfully embed functional and perfusable microchannels inside methacrylated gelatin (GelMA), star poly(ethylene glycol-co-lactide) acrylate (SPELA), poly(ethylene glycol) dimethacrylate (PEGDMA) and poly(ethylene glycol) diacrylate (PEGDA) hydrogels at different concentrations. In particular, GelMA hydrogels were used as a model to demonstrate the functionality of the fabricated vascular networks in improving mass transport, cellular viability and differentiation within the cell-laden tissue constructs. In addition, successful formation of endothelial monolayers within the fabricated channels was confirmed. Overall, our proposed strategy represents an effective technique for vascularization of hydrogel constructs with useful applications in tissue engineering and organs on a chip.

726 citations


Journal ArticleDOI
TL;DR: It is hoped that an improved fundamental and quantitative understanding of inertial fluid dynamic effects can lead to unprecedented capabilities to program fluid and particle flow towards automation of biomedicine, materials synthesis, and chemical process control.
Abstract: Microfluidics has experienced massive growth in the past two decades, and especially with advances in rapid prototyping researchers have explored a multitude of channel structures, fluid and particle mixtures, and integration with electrical and optical systems towards solving problems in healthcare, biological and chemical analysis, materials synthesis, and other emerging areas that can benefit from the scale, automation, or the unique physics of these systems. Inertial microfluidics, which relies on the unconventional use of fluid inertia in microfluidic platforms, is one of the emerging fields that make use of unique physical phenomena that are accessible in microscale patterned channels. Channel shapes that focus, concentrate, order, separate, transfer, and mix particles and fluids have been demonstrated, however physical underpinnings guiding these channel designs have been limited and much of the development has been based on experimentally-derived intuition. Here we aim to provide a deeper understanding of mechanisms and underlying physics in these systems which can lead to more effective and reliable designs with less iteration. To place the inertial effects into context we also discuss related fluid-induced forces present in particulate flows including forces due to non-Newtonian fluids, particle asymmetry, and particle deformability. We then highlight the inverse situation and describe the effect of the suspended particles acting on the fluid in a channel flow. Finally, we discuss the importance of structured channels, i.e. channels with boundary conditions that vary in the streamwise direction, and their potential as a means to achieve unprecedented three-dimensional control over fluid and particles in microchannels. Ultimately, we hope that an improved fundamental and quantitative understanding of inertial fluid dynamic effects can lead to unprecedented capabilities to program fluid and particle flow towards automation of biomedicine, materials synthesis, and chemical process control.

578 citations


Journal ArticleDOI
TL;DR: This work reports on a novel spiral microfluidic device with a trapezoidal cross-section for ultra-fast, label-free enrichment of CTCs from clinically relevant blood volumes, using the inherent Dean vortex flows present in curvilinear microchannels under continuous flow.
Abstract: The enumeration and characterization of circulating tumor cells (CTCs), found in the peripheral blood of cancer patients, provide a potentially accessible source for cancer diagnosis and prognosis. This work reports on a novel spiral microfluidic device with a trapezoidal cross-section for ultra-fast, label-free enrichment of CTCs from clinically relevant blood volumes. The technique utilizes the inherent Dean vortex flows present in curvilinear microchannels under continuous flow, along with inertial lift forces which focus larger CTCs against the inner wall. Using a trapezoidal cross-section as opposed to a traditional rectangular cross-section, the position of the Dean vortex core can be altered to achieve separation. Smaller hematologic components are trapped in the Dean vortices skewed towards the outer channel walls and eventually removed at the outer outlet, while the larger CTCs equilibrate near the inner channel wall and are collected from the inner outlet. By using a single spiral microchannel with one inlet and two outlets, we have successfully isolated and recovered more than 80% of the tested cancer cell line cells (MCF-7, T24 and MDA-MB-231) spiked in 7.5 mL of blood within 8 min with extremely high purity (400–680 WBCs mL−1; ∼4 log depletion of WBCs). Putative CTCs were detected and isolated from 100% of the patient samples (n = 10) with advanced stage metastatic breast and lung cancer using standard biomarkers (CK, CD45 and DAPI) with the frequencies ranging from 3–125 CTCs mL−1. We expect this simple and elegant approach can surmount the shortcomings of traditional affinity-based CTC isolation techniques as well as enable fundamental studies on CTCs to guide treatment and enhance patient care.

518 citations


Journal ArticleDOI
TL;DR: The ability of ExoChip to recover exosomes with intact RNA enabling profiling of exosomal-microRNAs through openarray analysis, which has potential applications in biomarker discovery.
Abstract: Membrane bound vesicles, including microvesicles and exosomes, are secreted by both normal and cancerous cells into the extracellular space and in blood circulation. These circulating extracellular vesicles (cirEVs) and exosomes in particular are recognized as a potential source of disease biomarkers. However, to exploit the use of circulatory exosomes as a biomarker, a rapid, high-throughput and reproducible method is required for their isolation and molecular analysis. We have developed a simple, low cost microfluidic-based platform to isolate cirEVs enriched in exosomes directly from blood serum allowing simultaneous capture and quantification of exosomes in a single device. To capture specific exosomes, we employed “ExoChip”, a microfluidic device fabricated in polydimethylsiloxane (PDMS) and functionalized with antibodies against CD63, an antigen commonly overexpressed in exosomes. Subsequent staining with a fluorescent carbocyanine dye (DiO) that specifically labels the exosomes, we quantitated exosomes using a standard plate-reader. Ten independent ExoChip experiments performed using serum obtained from five pancreatic cancer patients and five healthy individuals revealed a statistically significant increase (2.34 ± 0.31 fold, p < 0.001) in exosomes captured in cancer patients when compared to healthy individuals. Exosomal origins of ExoChip immobilized vesicles were further confirmed using immuno-electron-microscopy and Western blotting. In addition, we demonstrate the ability of ExoChip to recover exosomes with intact RNA enabling profiling of exosomal-microRNAs through openarray analysis, which has potential applications in biomarker discovery. Based on our findings, ExoChip is a well suited platform to be used as an exosome-based diagnostic and research tool for molecular screening of human cancers.

491 citations


Journal ArticleDOI
TL;DR: This Vortex approach offers significant advantages over existing technologies, especially in terms of processing time, sample concentration, applicability to various cancer types, cell integrity and purity, and widespread adoption by clinicians and biologists who desire to enumerate CTCs.
Abstract: A blood-based, low cost alternative to radiation intensive CT and PET imaging is critically needed for cancer prognosis and management of its treatment. "Liquid biopsies" of circulating tumor cells (CTCs) from a relatively non-invasive blood draw are particularly ideal, as they can be repeated regularly to provide up to date molecular information about the cancer, which would also open up key opportunities for personalized therapies. Beyond solely diagnostic applications, CTCs are also a subject of interest for drug development and cancer research. In this paper, we adapt a technology previously introduced, combining the use of micro-scale vortices and inertial focusing, specifically for the high-purity extraction of CTCs from blood samples. First, we systematically varied parameters including channel dimensions and flow rates to arrive at an optimal device for maximum trapping efficiency and purity. Second, we validated the final device for capture of cancer cell lines in blood, considering several factors, including the effect of blood dilution, red blood cell lysis and cell deformability, while demonstrating cell viability and independence on EpCAM expression. Finally, as a proof-of-concept, CTCs were successfully extracted and enumerated from the blood of patients with breast (N = 4, 25-51 CTCs per 7.5 mL) and lung cancer (N = 8, 23-317 CTCs per 7.5 mL). Importantly, samples were highly pure with limited leukocyte contamination (purity 57-94%). This Vortex approach offers significant advantages over existing technologies, especially in terms of processing time (20 min for 7.5 mL of whole blood), sample concentration (collecting cells in a small volume down to 300 μL), applicability to various cancer types, cell integrity and purity. We anticipate that its simplicity will aid widespread adoption by clinicians and biologists who desire to not only enumerate CTCs, but also uncover new CTC biology, such as unique gene mutations, vesicle secretion and roles in metastatic processes.

468 citations


Journal ArticleDOI
Mei He1, Jennifer Crow1, Marc Roth1, Yong Zeng1, Andrew K. Godwin1 
TL;DR: In this article, a microfluidic exosome analysis platform was proposed for non-invasive cancer diagnosis and monitoring of treatment response using plasma exosomes, which can be used to accelerate the exosomal analysis pipeline by integrating specific immunoisolation and targeted protein analysis.
Abstract: Developing blood-based tests is appealing for non-invasive disease diagnosis, especially when biopsy is difficult, costly, and sometimes not even an option. Tumor-derived exosomes have attracted increasing interest in non-invasive cancer diagnosis and monitoring of treatment response. However, the biology and clinical value of exosomes remains largely unknown due in part to current technical challenges in rapid isolation, molecular classification and comprehensive analysis of exosomes. Here we developed a new microfluidic approach to streamline and expedite the exosome analysis pipeline by integrating specific immunoisolation and targeted protein analysis of circulating exosomes. Compared to the conventional methods, our approach enables selective subpopulation isolation and quantitative detection of surface and intravesicular biomarkers directly from a minimally invasive amount of plasma samples (30 μL) within ~100 min with markedly improved detection sensitivity. Using this device, we demonstrated phenotyping of exosome subpopulations by targeting a panel of common exosomal and tumor-specific markers and multiparameter analyses of intravesicular biomarkers in the selected subpopulation. We were able to assess the total expression and phosphorylation levels of IGF-1R in non-small-cell lung cancer patients by probing plasma exosomes as a non-invasive alternative to conventional tissue biopsy. We foresee that the microfluidic exosome analysis platform will form the basis for critically needed infrastructures for advancing the biology and clinical utilization of exosomes.

368 citations


Journal ArticleDOI
TL;DR: Many interesting publications are collated to provide a comprehensive review of the development and diversity of this technology but also provide scope for future direction and detail the fundamentals for those wishing to design such devices for the first time.
Abstract: Deterministic lateral displacement (DLD), a hydrodynamic, microfluidic technology, was first reported by Huang et al. in 2004 to separate particles on the basis of size in continuous flow with a resolution of down to 10 nm. For 10 years, DLD has been extensively studied, employed and modified by researchers in terms of theory, design, microfabrication and application to develop newer, faster and more efficient tools for separation of millimetre, micrometre and even sub-micrometre sized particles. To extend the range of potential applications, the specific arrangement of geometric features in DLD has also been adapted and/or coupled with external forces (e.g. acoustic, electric, gravitational) to separate particles on the basis of other properties than size such as the shape, deformability and dielectric properties of particles. Furthermore, investigations into DLD performance where inertial and non-Newtonian effects are present have been conducted. However, the evolvement and application of DLD has not yet been reviewed. In this paper, we collate many interesting publications to provide a comprehensive review of the development and diversity of this technology but also provide scope for future direction and detail the fundamentals for those wishing to design such devices for the first time.

355 citations


Journal ArticleDOI
TL;DR: The resolution of an SL mail-order service and the main causes of resolution loss; the optical clarity of the devices and how to address the lack of clarity for imaging in the channels; and the future role that SL could play in the commercialization of microfluidic devices are evaluated.
Abstract: The vast majority of microfluidic devices are developed in PDMS by molding (“soft lithography”) because PDMS is an inexpensive material, has physicochemical properties that are well suited for biomedical and physical sciences applications, and design cycle lengths are generally adequate for prototype development. However, PDMS molding is tediously slow and thus cannot provide the high- or medium-volume production required for the commercialization of devices. While high-throughput plastic molding techniques (e.g. injection molding) exist, the exorbitant cost of the molds and/or the equipment can be a serious obstacle for device commercialization, especially for small startups. High-volume production is not required to reach niche markets such as clinical trials, biomedical research supplies, customized research equipment, and classroom projects. Crucially, both PDMS and plastic molding are layer-by-layer techniques where each layer is produced as a result of physicochemical processes not specified in the initial photomask(s) and where the final device requires assembly by bonding, all resulting in a cost that is very hard to predict at the start of the project. By contrast, stereolithography (SL) is an automated fabrication technique that allows for the production of quasi-arbitrary 3D shapes in a single polymeric material at medium-volume throughputs (ranging from a single part to hundreds of parts). Importantly, SL devices can be designed between several groups using CAD tools, conveniently ordered by mail, and their cost precisely predicted via a web interface. Here we evaluate the resolution of an SL mail-order service and the main causes of resolution loss; the optical clarity of the devices and how to address the lack of clarity for imaging in the channels; and the future role that SL could play in the commercialization of microfluidic devices.

314 citations


Journal ArticleDOI
TL;DR: Some of the emerging applications and the future opportunities and challenges created by the use of mobile phones and their embedded components for the development of next-generation imaging, sensing, diagnostics and measurement tools are discussed.
Abstract: In this article, I discuss some of the emerging applications and the future opportunities and challenges created by the use of mobile phones and their embedded components for the development of next-generation imaging, sensing, diagnostics and measurement tools. The massive volume of mobile phone users, which has now reached ~7 billion, drives the rapid improvements of the hardware, software and high-end imaging and sensing technologies embedded in our phones, transforming the mobile phone into a cost-effective and yet extremely powerful platform to run, e.g., biomedical tests, and perform scientific measurements that would normally require advanced laboratory instruments. This rapidly evolving and continuing trend will help us transform how medicine, engineering and sciences are practiced and taught globally.

314 citations


Journal ArticleDOI
TL;DR: A facile wettability patterning method is used to produce open microfluidic tracks that are capable of transporting a wide range of liquid volumes on-chip, overcoming viscous and other opposing forces at the pertinent length scales.
Abstract: Surface tension driven transport of liquids on open substrates offers an enabling tool for open micro total analysis systems that are becoming increasingly popular for low-cost biomedical diagnostic devices. The present study uses a facile wettability patterning method to produce open microfluidic tracks that – due to their shape, surface texture and chemistry – are capable of transporting a wide range of liquid volumes (~1–500 μL) on-chip, overcoming viscous and other opposing forces (e.g., gravity) at the pertinent length scales. Small volumes are handled as individual droplets, while larger volumes require repeated droplet transport. The concept is developed and demonstrated with coatings based on TiO2 filler particles, which, when present in adequate (~80 wt.%) quantities within a hydrophobic fluoroacrylic polymer matrix, form composites that are intrinsically superhydrophobic. Such composite coatings become superhydrophilic upon exposure to UV light (390 nm). A commercial laser printer-based photo-masking approach is used on the coating for spatially selective wettability conversion from superhydrophobic to superhydrophilic. Carefully designed wedge-patterned surface tension confined tracks on the open-air devices move liquid on them without power input, even when acting against gravity. Simple designs of wettability patterning are used on versatile substrates (e.g., metals, polymers, paper) to demonstrate complex droplet handling tasks, e.g., merging, splitting and metered dispensing, some of which occur in 3-D geometries. Fluid transport rates of up to 350 μL s−1 are attained. Applicability of the design on metal substrates allows these devices to be used also for other microscale engineering applications, e.g., water management in fuel cells.

281 citations


Journal ArticleDOI
TL;DR: The design considerations of representative microfluidic devices for rare cell isolation and analysis and a perspective on the development trends and promising research directions in this field are proposed.
Abstract: Rare cells are low-abundance cells in a much larger population of background cells. Conventional benchtop techniques have limited capabilities to isolate and analyze rare cells because of their generally low selectivity and significant sample loss. Recent rapid advances in microfluidics have been providing robust solutions to the challenges in the isolation and analysis of rare cells. In addition to the apparent performance enhancements resulting in higher efficiencies and sensitivity levels, microfluidics provides other advanced features such as simpler handling of small sample volumes and multiplexing capabilities for high-throughput processing. All of these advantages make microfluidics an excellent platform to deal with the transport, isolation, and analysis of rare cells. Various cellular biomarkers, including physical properties, dielectric properties, as well as immunoaffinities, have been explored for isolating rare cells. In this Focus article, we discuss the design considerations of representative microfluidic devices for rare cell isolation and analysis. Examples from recently published works are discussed to highlight the advantages and limitations of the different techniques. Various applications of these techniques are then introduced. Finally, a perspective on the development trends and promising research directions in this field are proposed.

Journal ArticleDOI
TL;DR: The current development of microfluidic-based spinning techniques for producing micro- and nano-scale fibers based on different solidification methods, platforms, geometries, or biomaterials are described and the emerging applications of fibers as bottom-up scaffolds such as cell encapsulation or guidance are highlighted.
Abstract: Microfluidic technologies have recently been shown to hold significant potential as novel tools for producing micro- and nano-scale structures for a variety of applications in tissue engineering and cell biology. Over the last decade, microfluidic spinning has emerged as an advanced method for fabricating fibers with diverse shapes and sizes without the use of complicated devices or facilities. In this critical review, we describe the current development of microfluidic-based spinning techniques for producing micro- and nano-scale fibers based on different solidification methods, platforms, geometries, or biomaterials. We also highlight the emerging applications of fibers as bottom-up scaffolds such as cell encapsulation or guidance for use in tissue engineering research and clinical practice.

Journal ArticleDOI
TL;DR: The results reported here verify the reproducible and transferable nature of using 3D printing as a fabrication technique, as devices and electrodes were moved between labs multiple times during completion of the study.
Abstract: We report two 3D printed devices that can be used for electrochemical detection In both cases, the electrode is housed in commercially available, polymer-based fittings so that the various electrode materials (platinum, platinum black, carbon, gold, silver) can be easily added to a threaded receiving port printed on the device; this enables a module-like approach to the experimental design, where the electrodes are removable and can be easily repolished for reuse after exposure to biological samples The first printed device represents a microfluidic platform with a 500 × 500 μm channel and a threaded receiving port to allow integration of either polyetheretherketone (PEEK) nut-encased glassy carbon or platinum black (Pt-black) electrodes for dopamine and nitric oxide (NO) detection, respectively The embedded 1 mm glassy carbon electrode had a limit of detection (LOD) of 500 nM for dopamine and a linear response (R2 = 099) for concentrations between 25–500 μM When the glassy carbon electrode was coated with 005% Nafion, significant exclusion of nitrite was observed when compared to signal obtained from equimolar injections of dopamine When using flow injection analysis with a Pt/Pt-black electrode and standards derived from NO gas, a linear correlation (R2 = 099) over a wide range of concentrations (76–190 μM) was obtained, with the LOD for NO being 1 μM The second application showcases a 3D printed fluidic device that allows collection of the biologically relevant analyte adenosine triphosphate (ATP) while simultaneously measuring the release stimulus (reduced oxygen concentration) The hypoxic sample (48 ± 05 ppm oxygen) released 24 ± 04 times more ATP than the normoxic sample (84 ± 06 ppm oxygen) Importantly, the results reported here verify the reproducible and transferable nature of using 3D printing as a fabrication technique, as devices and electrodes were moved between labs multiple times during completion of the study

Journal ArticleDOI
TL;DR: The development of a geometrically enhanced mixing (GEM) chip for high-efficiency and high-purity tumor cell capture and the release and culture of the captured tumor cells, as well as the isolation of CTCs from cancer patients are reported.
Abstract: Circulating tumor cells (CTCs) from peripheral blood hold important information for cancer diagnosis and disease monitoring. Analysis of this “liquid biopsy” holds the promise to usher in a new era of personalized therapeutic treatments and real-time monitoring for cancer patients. But the extreme rarity of CTCs in blood makes their isolation and characterization technologically challenging. This paper reports the development of a geometrically enhanced mixing (GEM) chip for high-efficiency and high-purity tumor cell capture. We also successfully demonstrated the release and culture of the captured tumor cells, as well as the isolation of CTCs from cancer patients. The high-performance microchip is based on geometrically optimized micromixer structures, which enhance the transverse flow and flow folding, maximizing the interaction between CTCs and antibody-coated surfaces. With the optimized channel geometry and flow rate, the capture efficiency reached >90% with a purity of >84% when capturing spiked tumor cells in buffer. The system was further validated by isolating a wide range of spiked tumor cells (50–50 000) in 1 mL of lysed blood and whole blood. With the combination of trypsinization and high flow rate washing, captured tumor cells were efficiently released. The released cells were viable and able to proliferate, and showed no difference compared with intact cells that were not subjected to the capture and release process. Furthermore, we applied the device for detecting CTCs from metastatic pancreatic cancer patients' blood; and CTCs were found from 17 out of 18 samples (>94%). We also tested the potential utility of the device in monitoring the response to anti-cancer drug treatment in pancreatic cancer patients, and the CTC numbers correlated with the clinical computed tomograms (CT scans) of tumors. The presented technology shows great promise for accurate CTC enumeration, biological studies of CTCs and cancer metastasis, as well as for cancer diagnosis and treatment monitoring.

Journal ArticleDOI
TL;DR: This review focuses on efforts to combine and integrate different magnetically actuated assay steps, with the vision that it will become possible in the future to realize integrated lab-on-chip biosensing assays in which all assay process steps are controlled and optimized by magnetic forces.
Abstract: The demand for easy to use and cost effective medical technologies inspires scientists to develop innovative lab-on-chip technologies for point-of-care in vitro diagnostic testing. To fulfill medical needs, the tests should be rapid, sensitive, quantitative, and miniaturizable, and need to integrate all steps from sample-in to result-out. Here, we review the use of magnetic particles actuated by magnetic fields to perform the different process steps that are required for integrated lab-on-chip diagnostic assays. We discuss the use of magnetic particles to mix fluids, to capture specific analytes, to concentrate analytes, to transfer analytes from one solution to another, to label analytes, to perform stringency and washing steps, and to probe biophysical properties of the analytes, distinguishing methodologies with fluid flow and without fluid flow (stationary microfluidics). Our review focuses on efforts to combine and integrate different magnetically actuated assay steps, with the vision that it will become possible in the future to realize integrated lab-on-chip biosensing assays in which all assay process steps are controlled and optimized by magnetic forces.

Journal ArticleDOI
TL;DR: Current methods for the enrichment and detection of circulating tumor cells are summarized and the key challenges and perspectives of CTC analyses within the context of improved clinical management of cancer patients are discussed.
Abstract: Hematogeneous tumor cell dissemination is a key step in cancer progression. The detection of CTCs in the peripheral blood of patients with solid epithelial tumors (e.g., breast, prostate, lung and colon cancer) holds great promise, and many exciting technologies have been developed over the past years. However, the detection and molecular characterization of circulating tumor cells (CTCs) remain technically challenging. The identification and characterization of CTCs require extremely sensitive and specific analytical methods, which are usually a combination of complex enrichment and detection procedures. CTCs occur at very low concentrations of one tumor cell in the background of millions of normal blood cells and the epithelial–mesenchymal plasticity of CTCs can hamper their detection by the epithelial markers used in current CTC assays. In the present review, we summarize current methods for the enrichment and detection of CTCs and discuss the key challenges and perspectives of CTC analyses within the context of improved clinical management of cancer patients.

Journal ArticleDOI
TL;DR: The fabrication of conventional PDMS on glass lab-on-a-chip (LOC) devices, using templates printed with a commercial micro-stereo lithography 3D printer, is demonstrated, which promotes broader access to established LOC configurations with minimal fabrication requirements, relieves LOC fabrication from design skills and provides a versatile LOC development platform.
Abstract: The fabrication of conventional PDMS on glass lab-on-a-chip (LOC) devices, using templates printed with a commercial (2299 US$) micro-stereo lithography 3D printer, is demonstrated. Printed templates replace clean room and photolithographic fabrication resources and deliver resolutions of 50 μm, and up to 10 μm in localized hindrances, whereas the templates are smooth enough to allow direct transfer and proper sealing to glass substrates. 3D printed templates accommodate multiple thicknesses, from 50 μm up to several mm within the same template, with no additional processing cost or effort. This capability is exploited to integrate silicone tubing easily, to improve micromixer performance and to produce multilevel fluidics with simple access to independent functional surfaces, which is illustrated by time-resolved glucose detection. The templates are reusable, can be fabricated in under 20 min, with an average cost of 0.48 US$, which promotes broader access to established LOC configurations with minimal fabrication requirements, relieves LOC fabrication from design skills and provides a versatile LOC development platform.

Journal ArticleDOI
TL;DR: A versatile microfluidic fluorescence-activated cell sorter that uses acoustic actuation to sort cells or drops at ultra-high rates and enables a rich variety of more sophisticated applications.
Abstract: We describe a versatile microfluidic fluorescence-activated cell sorter that uses acoustic actuation to sort cells or drops at ultra-high rates. Our acoustic sorter combines the advantages of traditional fluorescence-activated cell (FACS) and droplet sorting (FADS) and is applicable for a multitude of objects. We sort aqueous droplets, at rates as high as several kHz, into two or even more outlet channels. We can also sort cells directly from the medium without prior encapsulation into drops; we demonstrate this by sorting fluorescently labeled mouse melanoma cells in a single phase fluid. Our acoustic microfluidic FACS is compatible with standard cell sorting cytometers, yet, at the same time, enables a rich variety of more sophisticated applications.

Journal ArticleDOI
TL;DR: A novel, multiparametric microphysiometry system for the dynamic online monitoring of human cancer cell metabolism and the drug screening application was demonstrated by detecting alteration and recovery effects of cellular metabolism induced by the addition of substances to the medium.
Abstract: We present a novel, multiparametric microphysiometry system for the dynamic online monitoring of human cancer cell metabolism. The optically transparent, modular, hybrid microsystem is based on a glass chip and combines a cell cultivation chamber, microfluidics and metabolic monitoring with fully integrated chemo- and biosensors. pH and oxygen are measured in the cell culture area, and biosensors for lactate and glucose are connected downstream by microfluidics. The wafer-level fabrication features thin-film platinum and iridium oxide microelectrodes on a glass chip, microfluidics in an epoxy resist, a hybrid assembly and an on-chip reference electrode. The reliable analytical performance of the sensors in cell culture medium was demonstrated. The pH sensors exhibit a long-term stable, linear response. The oxygen sensors show a linear behaviour, which is also observed for low oxygen concentrations. Glucose and lactate measurements show a linear, long-term stable, selective and reversible behaviour in the desired range. T98G human brain cancer cells were cultivated and cell culture metabolism was measured on-chip. Stop/flow cycles were applied and extracellular acidification, respiration, glucose consumption and lactate production were quantified. Long-term metabolic rates were determined and all parameters could be measured in the outlet channel. A placement downstream of the cell cultivation area for biosensors was realised. A highly effective medium exchange and undiluted sampling from the cell culture chamber with low flow rates (2 μl min−1) and low volumes (15 μl per cycle) were achieved. The drug screening application was demonstrated by detecting alteration and recovery effects of cellular metabolism induced by the addition of substances to the medium.

Journal ArticleDOI
TL;DR: Foam mobility control and novel oil displacement mechanisms were observed in a microfluidic device representing a porous media system with layered permeability, suggesting that it is not appropriate to treat foam as a homogeneous dispersion of gas and liquid.
Abstract: Foam mobility control and novel oil displacement mechanisms were observed in a microfluidic device representing a porous media system with layered permeability. Foam was pre-generated using a flow-focusing microfluidic device and injected into an oil-wet, oil-saturated 2-D PDMS microfluidic device. The device is designed with a central fracture flanked by high-permeability and low-permeability zones stratified in the direction of injection. A 1 : 1, 1% blend of alpha olefin sulfonate 14–16 (AOS) and lauryl betaine (LB) surfactants produced stable foam in the presence of paraffin oil. The oil saturation and pressure drop across the microfluidic device were measured as a function of time and the injected pore volume, indicating an increase in apparent viscosity for foam with an accompanying decrease in oil saturation. In contrast to the control experiments, foam was shown to more effectively mobilize trapped oil by increasing the flow resistance in the fracture and high-permeability zones and by diverting the surfactant solution into adjacent low-permeability zones. The foam was observed to separate into gas-rich and aqueous-rich phases depending on matrix permeability, suggesting that it is not appropriate to treat foam as a homogeneous dispersion of gas and liquid.

Journal ArticleDOI
TL;DR: Using the 'GI tract-liver-other tissues' system allowed us to observe compounding effects and detect liver tissue injury at lower nanoparticle concentrations than was expected from experiments with single tissues, and indicates that body-on-a-chip devices are highly relevant in vitro models for evaluating nanoparticle interactions with human tissues.
Abstract: The use of nanoparticles in medical applications is highly anticipated, and at the same time little is known about how these nanoparticles affect human tissues. Here we have simulated the oral uptake of 50 nm carboxylated polystyrene nanoparticles with a microscale body-on-a-chip system (also referred to as multi-tissue microphysiological system or micro Cell Culture Analog). Using the ‘GI tract–liver–other tissues’ system allowed us to observe compounding effects and detect liver tissue injury at lower nanoparticle concentrations than was expected from experiments with single tissues. To construct this system, we combined in vitro models of the human intestinal epithelium, represented by a co-culture of enterocytes (Caco-2) and mucin-producing cells (TH29-MTX), and the liver, represented by HepG2/C3A cells, within one microfluidic device. The device also contained chambers that together represented the liquid portions of all other organs of the human body. Measuring the transport of 50 nm carboxylated polystyrene nanoparticles across the Caco-2/HT29-MTX co-culture, we found that this multi-cell layer presents an effective barrier to 90.5 ± 2.9% of the nanoparticles. Further, our simulation suggests that a larger fraction of the 9.5 ± 2.9% nanoparticles that travelled across the Caco-2/HT29-MTX cell layer were not large nanoparticle aggregates, but primarily single nanoparticles and small aggregates. After crossing the GI tract epithelium, nanoparticles that were administered in high doses estimated in terms of possible daily human consumption (240 and 480 × 1011 nanoparticles mL−1) induced the release of aspartate aminotransferase (AST), an intracellular enzyme of the liver that indicates liver cell injury. Our results indicate that body-on-a-chip devices are highly relevant in vitro models for evaluating nanoparticle interactions with human tissues.

Journal ArticleDOI
TL;DR: Through a series of human trials, the system can accurately quantify total cholesterol levels in blood within 60 s by imaging standard test strips and is optimized to improve measurement sensitivity and reproducibility across different individual smartphones.
Abstract: Home self-diagnostic tools for blood cholesterol monitoring have been around for over a decade but their widespread adoption has been limited by the relatively high cost of acquiring a quantitative test-strip reader, complicated procedure for operating the device, and inability to easily store and process results. To address this we have developed a smartphone accessory and software application that allows for the quantification of cholesterol levels in blood. Through a series of human trials we demonstrate that the system can accurately quantify total cholesterol levels in blood within 60 s by imaging standard test strips. In addition, we demonstrate how our accessory is optimized to improve measurement sensitivity and reproducibility across different individual smartphones. With the widespread adoption of smartphones and increasingly sophisticated image processing technology, accessories such as the one presented here will allow cholesterol monitoring to become more accurate and widespread, greatly improving preventive care for cardiovascular disease.

Journal ArticleDOI
TL;DR: Two bioprinting approaches are discussed in detail - projection printing and laser-based systems - where each method is capable of modulating multiple scaffold parameters, such as 3D architecture, mechanical properties, and porosity.
Abstract: Light-assisted 3D direct-printing of biomaterials and cellular-scaffolds has the potential to develop novel lab-on-a-chip devices (LOCs) for a variety of biomedical applications, from drug discovery and diagnostic testing to in vitro tissue engineering and regeneration. Direct-writing describes a broad family of fabrication methods that typically employ computer-controlled translational stages to manufacture structures at multi-length scales. This review focuses on light-assisted direct-write fabrication for generating 3D functional scaffolds with precise micro- and nano-architecture, using both synthetic as well as naturally derived biomaterials. Two bioprinting approaches are discussed in detail – projection printing and laser-based systems – where each method is capable of modulating multiple scaffold parameters, such as 3D architecture, mechanical properties (e.g. stiffness), Poisson's ratio, chemical gradients, biological cell distributions, and porosity. The light-assisted direct-writing techniques described in this review provide the reader with alternative approaches to fabricate 3D biomaterials for utility in LOCs.

Journal ArticleDOI
TL;DR: A high-throughput method for single cell screening by microfluidic droplet sorting is applied to a whole-genome mutated yeast cell library yielding improved production hosts of secreted industrial enzymes.
Abstract: A high-throughput method for single cell screening by microfluidic droplet sorting is applied to a whole-genome mutated yeast cell library yielding improved production hosts of secreted industrial enzymes. The sorting method is validated by enriching a yeast strain 14 times based on its α-amylase production, close to the theoretical maximum enrichment. Furthermore, a 105 member yeast cell library is screened yielding a clone with a more than 2-fold increase in α-amylase production. The increase in enzyme production results from an improvement of the cellular functions of the production host in contrast to previous droplet-based directed evolution that has focused on improving enzyme protein structure. In the workflow presented, enzyme producing single cells are encapsulated in 20 pL droplets with a fluorogenic reporter substrate. The coupling of a desired phenotype (secreted enzyme concentration) with the genotype (contained in the cell) inside a droplet enables selection of single cells with improved enzyme production capacity by droplet sorting. The platform has a throughput over 300 times higher than that of the current industry standard, an automated microtiter plate screening system. At the same time, reagent consumption for a screening experiment is decreased a million fold, greatly reducing the costs of evolutionary engineering of production strains.

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TL;DR: This report specifically focuses on development of a practical app for immediate point-of-care (POC) multi-analyte sensing without additional devices that digitizes the colours of a colorimetric sensor array.
Abstract: Here we report development of a smartphone app (application) that digitizes the colours of a colorimetric sensor array. A conventional colorimetric sensor array consists of multiple paper-based sensors, and reports the detection results in terms of colour change. Evaluation of the colour changes is normally done by the naked eye, which may cause uncertainties due to personal subjectivity and the surrounding conditions. Solutions have been particularly sought in smartphones as they are capable of spectrometric functions. Our report specifically focuses on development of a practical app for immediate point-of-care (POC) multi-analyte sensing without additional devices. First, the individual positions of the sensors are automatically identified by the smartphone; second, the colours measured at each sensor are digitized based on a correction algorithm; and third, the corrected colours are converted to concentration values by pre-loaded calibration curves. All through these sequential processes, the sensor array taken in a smartphone snapshot undergoes laboratory-level spectrometry. The advantages of inexpensive and convenient paper-based colorimetry and the ubiquitous smartphone are tied to achieve a ready-to-go POC diagnosis.

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TL;DR: This review reviews biophysical label-free technologies that have been developed for CTC separation, including techniques based on filtration, hydrodynamic chromatography, and dielectrophoresis and discusses requirements for subsequent characterization of CTCs.
Abstract: Circulating tumor cells (CTCs) are malignant cells shed into the bloodstream from a tumor that have the potential to establish metastases in different anatomical sites. The separation and subsequent characterization of these cells is emerging as an important tool for both biomarker discovery and the elucidation of mechanisms of metastasis. Established methods for separating CTCs rely on biochemical markers of epithelial cells that are known to be unreliable because of epithelial-to-mesenchymal transition, which reduces expression for epithelial markers. Emerging label-free separation methods based on the biophysical and biomechanical properties of CTCs have the potential to address this key shortcoming and present greater flexibility in the subsequent characterization of these cells. In this review we first present what is known about the biophysical and biomechanical properties of CTCs from historical studies and recent research. We then review biophysical label-free technologies that have been developed for CTC separation, including techniques based on filtration, hydrodynamic chromatography, and dielectrophoresis. Finally, we evaluate these separation methods and discuss requirements for subsequent characterization of CTCs.

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TL;DR: The major developments in systems for flow chemistry are reviewed and limitations underlying the development of chip-scale integrated systems are discussed.
Abstract: Chemical synthesis in microsystems has evolved from simple proof-of-principle examples to become a general technique in academia and industry. Numerous such “flow chemistry” applications are now found in pharmaceutical and fine chemical synthesis. Much of the development has been based on systems employing macroscopic flow components and tubes, rather than the integrated chip technology envisioned by the lab-on-a-chip community. We review the major developments in systems for flow chemistry and discuss limitations underlying the development of chip-scale integrated systems.

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TL;DR: A new strategy is presented, in which FLAE is combined with femtosecond laser two-photon polymerization to realize a new type of biochip termed the ship-in-a-bottle biochip.
Abstract: Femtosecond lasers have unique characteristics of ultrashort pulse width and extremely high peak intensity; however, one of the most important features of femtosecond laser processing is that strong absorption can be induced only at the focus position inside transparent materials due to nonlinear multiphoton absorption. This exclusive feature makes it possible to directly fabricate three-dimensional (3D) microfluidic devices in glass microchips by two methods: 3D internal modification using direct femtosecond laser writing followed by chemical wet etching (femtosecond laser-assisted etching, FLAE) and direct ablation of glass in water (water-assisted femtosecond laser drilling, WAFLD). Direct femtosecond laser writing also enables the integration of micromechanical, microelectronic, and microoptical components into the 3D microfluidic devices without stacking or bonding substrates. This paper gives a comprehensive review on the state-of-the-art femtosecond laser 3D micromachining for the fabrication of microfluidic, optofluidic, and electrofluidic devices. A new strategy (hybrid femtosecond laser processing) is also presented, in which FLAE is combined with femtosecond laser two-photon polymerization to realize a new type of biochip termed the ship-in-a-bottle biochip.

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TL;DR: The functions and uses of a modern flow cytometer are detailed, the recent and historical contributions of microfluidics and microfabricated devices to field of flow cytometry are reviewed, current application areas are examined, and opportunities for the synergistic application of microFabrication approaches are suggested.
Abstract: A modern flow cytometer can analyze and sort particles on a one by one basis at rates of 50 000 particles per second. Flow cytometers can also measure as many as 17 channels of fluorescence, several angles of scattered light, and other non-optical parameters such as particle impedance. More specialized flow cytometers can provide even greater analysis power, such as single molecule detection, imaging, and full spectral collection, at reduced rates. These capabilities have made flow cytometers an invaluable tool for numerous applications including cellular immunophenotyping, CD4+ T-cell counting, multiplex microsphere analysis, high-throughput screening, and rare cell analysis and sorting. Many bio-analytical techniques have been influenced by the advent of microfluidics as a component in analytical tools and flow cytometry is no exception. Here we detail the functions and uses of a modern flow cytometer, review the recent and historical contributions of microfluidics and microfabricated devices to field of flow cytometry, examine current application areas, and suggest opportunities for the synergistic application of microfabrication approaches to modern flow cytometry.

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TL;DR: It is believed that the widespread availability of smartphone technology and the capabilities they offer in terms of computation, communication, social networking, and imaging will be transformative to the deployment of lab-on-a-chip type technology both in the developed and developing world.
Abstract: The rapid expansion of mobile technology is transforming the biomedical landscape. By 2016 there will be 260 M active smartphones in the US and millions of health accessories and software “apps” running off them. In parallel with this have come major technical achievements in lab-on-a-chip technology leading to incredible new biochemical sensors and molecular diagnostic devices. Despite these advancements, the uptake of lab-on-a-chip technologies at the consumer level has been somewhat limited. We believe that the widespread availability of smartphone technology and the capabilities they offer in terms of computation, communication, social networking, and imaging will be transformative to the deployment of lab-on-a-chip type technology both in the developed and developing world. In this paper we outline why we believe this is the case, the new business models that may emerge, and detail some specific application areas in which this synergy will have long term impact, namely: nutrition monitoring and disease diagnostics in limited resource settings.